The project in progress has as its goal, the understanding of the role of proteins containing the vitamin K-dependent, calcium binding amino acid, gamma-carboxyglutamic Acid (Gla), in the development of calcific cardiovascular disease. The disease states currently under study are: atherosclerosis, calcific aortic stenosis, porcine xenograft valve degeneration, and the mineralization of left ventricular assist device pump bladder membranes. Progress to date has revealed protein bound Gla to occur in all of the above mentioned calcific cardiovascular diseases, and the data thus far indicate that Gla content appears to be related to pathologic severity, and tissue calcium levels. A unique Gla containing protein, which has been named atherocalcin, has been isolated from human atherosclerotic aorta and is currently being characterized. In addition, Gla containing proteins have been shown to be present in animal atherosclerotic lesions (both pigeon and nonhuman primate tissue), and in porcine xenograft valve calcification, occurring in calf implants, thereby establishing relevant animal model systems with which to develop an approach to the understanding of Gla containing proteins and their function in the human disorders under study. Studies in progress will further probe the importance of Gla containing proteins in the pathogenesis of calcific cardiovascular diseases. Continued isolation and characterization of atherocalcin (and any other Gla containing proteins occurring in atherosclerosis) will be carried out, and efforts will be made to obtain an antiserum to atherocalcin in order to develop immunoassays including: a radioimmunoassay; and an immunofluorescent procedure. The effects of warfarin on the development of atherosclerosis in White Carneau Pigeons and on porcine xenograft valve mineralization in calves will be examined to study the effects of vitamin K-antagonism on these disease processes. Finally, organ culture incubations with fresh atherosclerotic tissue, and freshly removed valve specimens (both xenograft, and calcific aortic stenosis) will be carried out to study the determinants of in situ biosynthesis of Gla containing proteins.